JPH0624598B2 - Blood purification material - Google Patents

Description

TECHNICAL FIELD The present invention relates to a plasma purification material used for removing harmful metals such as aluminum and pathogenic factors such as hepatitis B antigen from plasma.

[Prior Art and its Problems] Rheumatoid arthritis, myasthenia gravis, systemic lupus erythematosus (SLE), severe hepatitis, post-transfusion purpura, high-viscosity syndrome, Lefsum disease, leukemia, Good Bascher syndrome,
Due to thrombotic thrombocytopenic purpura, blood group mismatch pregnancy, sickle cell anemia, rapidly progressive glomerulonephritis associated with vasculitis, chronic liver failure, thrombocytosis, cryoglobulinemia, multiple myeloma Intractable diseases / ordinary diseases such as renal insufficiency, cancer, etc. have not been well understood in the cause and treatment method.

However, in these intractable diseases and odd diseases, the concentration of pathogenic factors such as immunoglobulin, IgG, immune complex, rheumatoid factor, and immunosuppressive factor in blood or body fluid is increased, and these pathogenic factors are removed from blood. In recent years, it has become clear that doing so can improve the symptoms.

Therefore, plasmapheresis is being widely used as a treatment for removing pathogenic factors.

Plasma exchange therapy is a treatment method in which plasma of a patient containing pathogenic factors and the like is replaced with plasma that does not contain these factors.

This treatment method requires 2 to 4 plasma at a time, but there is a problem in that it is difficult to continuously obtain such a large amount of plasma. In addition, there is a problem that the probability of infection with hepatitis and the like increases when plasma exchange is performed.

As a therapeutic method for solving such a problem, various methods have been investigated in which only plasma is separated from the blood of the patient, the plasma is purified, and then returned to the blood of the patient again.

One of the methods is a method of filtering plasma using a porous hollow membrane of an organic polymer compound. However, since this method screens plasma components only by the size of the molecule, it may remove necessary components. Further, there is a problem that the removal component fluctuates due to the clogging of the film.

On the other hand, various blood purification methods for selectively removing only specific unnecessary components using an adsorbent have been investigated, and some of them have already been clinically applied. Examples of these adsorbents include protein A-supporting adsorbents, DNA-supporting adsorbents, tryptophan-supporting adsorbents, methylated albumin-supporting adsorbents, heparin-supporting adsorbents, and other adsorbents that carry antigens and antibodies. An adsorbent in which a polymer is supported as a ligand using a biological compound is used.

Although these adsorbents using a bio-derived compound are excellent in adsorption specificity, they have a problem in that the adsorption characteristics are deteriorated during production, sterilization, and storage. Further, particularly in the case of an adsorbent having an antigen or an antibody as a ligand, the antigen or the antibody separated from the adsorbent may form an immune complex in the patient's body and cause a side effect.

Further, in recent years, the number of artificial dialysis patients has been increasing year by year in Japan, and it is said that the number is about 70,000 at present. Among them, particularly in patients undergoing dialysis for a long period of time, there is a problem that the amount of aluminum in the blood is significantly increased and the bone is weakened. As a countermeasure against this, for example, oral medication has been administered, but it has little effect.

[Technical Means for Solving Problems] The present invention relates to a blood purification material comprising a polymer having a side chain with a functional group selected from the group consisting of an amidoxime group, a 2-oxahydrazide group, and an acid hydrazide group.

The polymer constituting the blood purification material of the present invention can be synthesized, for example, according to the following method.

(1) A method for synthesizing a polymer having an amidoxime group in its side chain.

It can be synthesized by reacting a polymer having a nitrile group in the side chain with hydroxylamine.

As the polymer having a nitrile group in the side chain, an acrylonitrile homopolymer or a copolymer with another vinyl compound can be used.

Other vinyl compounds include divinylbenzene, vinyl acetate, styrene, vinylpyrrolidone, N-vinylacetamide, acrylic acid, methacrylic acid, acrylamide,
Vinyl chloride, vinylidene chloride, methacrylonitrile, methyl vinyl ketone, vinyl pyridine, diethyl maleate, fumaronitrile, diethyl fumarate, methyl triethylene glycol dimethacrylate, divinyl ethylene glycol, ethylene, propylene, butylene, butadiene, and isoprene. be able to.

The acrylonitrile polymer can be amidoxime-ized by reacting it with hydroxyamine hydrochloride and sodium hydroxide in a solvent at 50 to 100 ° C. As the solvent, polar solvents such as water, various alcohols and aprotic polar solvents can be used.

(2) A method for synthesizing a polymer having a 2-oxalohydroxycarbonyl group in its side chain.

(a) 1-methacryloyl-2-oxalohydrazine or 1
-Acroyl-2-oxalohydrazine can be synthesized alone or by a method of copolymerizing with another vinyl compound. As other vinyl compounds, those described in (1) and acrylonitrile can be used. The polymerization can be carried out under usual conditions and methods.

(b) It can also be synthesized by a method of hydrolyzing a polymer compound obtained by homopolymerization of the vinyl compound represented by the general formula (I) or copolymerization with another vinyl compound.

Examples of the vinyl compound represented by the general formula (I) include 1-methacroyl-2-methoxallylhydrazine, 1-methacroyl-2-ethoxalylhydrazine, 1-methacroyl-2-heptoxalylhydrazine, 1-methacroyl-2- Dodecoxalylhydrazine, 1-Methacloyl-2-octadecoxalylhydrazine, 1-Acroyl-2-methoxalylhydrazine, 1-Acroyl-2-ethoxalylhydrazine, 1-Acroyl-2-heptoxalylhydrazine, 1 -Acroyl-2-dodecoxalylhydrazine, 1-acroyl-2-octadecoxalylhydrazine and the like can be mentioned.

As the other vinyl compound, those mentioned in (2) can be used.

The polymerization can be carried out by a method usually used for polymerizing vinyl compounds.

It can also be synthesized by the reaction of (c) a polymer compound containing an acid chloride group, an acid amide group, or an acid anhydride group with oxalohydrazine.

The polymer compound containing an acid chloride group is obtained by converting a polymer compound containing a carboxyl group into phosgene, phosphorus pentoxide,
It can be synthesized by a method of reacting with a chlorinating reagent such as thionyl chloride, or by polymerizing a vinyl compound containing an acid chloride group. As the polymer compound containing a carboxyl group, acrylic acid, methacrylic acid,
Polymers of p-vinylbenzoic acid or copolymers of these compounds with the vinyl compound (1) are particularly preferable.

Further, as the vinyl compound containing an acid chloride group,
Acrylic acid chloride, methacrylic acid chloride, p-
Vinyl benzoyl chloride or the like can be used.

As the polymer compound containing an acid anhydride group, a copolymer of maleic anhydride or the like can be generally used.

The polymer compound having an acid amide group can be synthesized by a method of homopolymerizing or copolymerizing a vinyl compound having an acid amide group or a method of heating and dehydrating an ammonium salt of a polymer compound having a carboxyl group.

The reaction of the polymer compound containing an acid chloride group, an acid amide group, or an acid anhydride group with oxalohydrazine is preferably performed in an organic solvent. As the organic solvent, N-
Methyl-2-pyrrolidone, dimethylformamide, dimethylacetamide, benzene, toluene, xylene, dioxane and the like can be preferably used. The reaction is carried out at −10 ° C. to 150 ° C., preferably 10 to 80 ° C. for a polymer compound having an acid chloride. In the case of the polymer compound having an acid anhydride group, 20 to 150 ° C. is preferable as the reaction temperature. In the case of a polymer compound having an acid amide group, it is usually performed at 100 ° C to 250 ° C, preferably 150 ° C to 220 ° C.

When the acid chloride groups are close to each other in the polymer compound, or when the acid chloride groups are acid anhydride groups, an imide bond may be formed by the reaction with oxalohydrazine. In such a case, the desired 2-oxalohydrazinocarbonyl group can be obtained by treating the reaction product with dilute alkaline water.

(3) A method for synthesizing a polymer compound containing an acid hydrazide group.

It can be synthesized by the reaction of a polymer compound containing a carboxylic acid ester group or an acid chloride group with human lazine.

Examples of polymer compounds used for such purposes include:
Homopolymers of unsaturated ester compounds such as methyl acrylate and methyl methacrylate or copolymers with vinyl compounds described in (1), or esterified products of natural polymer compounds such as alginic acid and pectic acid, or It is preferable to use acid chlorides of these polymer compounds.

The hydrazine may be anhydrous hydrazine or hydrazine hydrate, but hydrazine hydrate is preferred.

The reaction temperature is preferably 50 to 200 ° C.

The shape of the blood purification material composed of these polymer compounds is preferably fibrous or particulate for easy handling. In the case of particles, the diameter is preferably 10 to 5000 μm, more preferably 50 to 1000 μm in view of clogging when packed in a column and adsorption rate.

As a method of purifying blood using the blood purification material of the present invention, the following method is preferable.

Plasma separated from whole blood by centrifugation or membrane separation is passed through a column filled with the blood purification agent of the present invention, or this plasma and the blood purification agent of the present invention are put in a container and stirred for a certain period of time. The blood purification agent is removed by filtration or the like.

Purification of blood is 10-42 ℃, preferably 25-37 ℃
It is preferable to carry out.

[Operation of the Present Invention] The blood purification material of the present invention is an unwanted substance such as aluminum in the blood, immunoglobulin, pathogenic factor IgG of multiple myeloma, immune complex, rheumatoid factor, or rejection factor of an organ transplant patient. Adsorbs well.

Also, anti-DNA in plasma of patients with systemic lupus erythematosus
Antibodies such as antibodies, immune complexes, anti-cycloglobulin antibodies and anti-thyroid microsome antibodies in the plasma of patients with Hashimoto's disease are also well adsorbed.

Immunosuppressive factors generated in the blood of cancer patients, EIDS viruses (HIV) in the blood of EIDS patients, hepatitis B viruses, etc. are also well adsorbed.

[Effects of the Present Invention] In the plasma purification method using the blood purification material of the present invention, unlike the method using a porous hollow fiber membrane of an organic polymer, problems such as clogging hardly occur. Also, unlike the method using an adsorbent carrying an antigen or an antibody, manufacturing, sterilization,
There is no problem of causing deterioration of adsorption characteristics during storage,
There is almost no risk that the antigen or antibody desorbed from the adsorbent will form an immune complex in the patient's body and cause side effects.

[Examples] Examples of the present invention will be shown below.

(1) Production of blood purification material having amidoxime group Example 1 (a) Synthesis of acrylonitrile copolymer 51.9 g of acrylonitrile, 62.3 g of divinylbenzene (purity 55%, removal of polymerization inhibitor), 80 ml of toluene , And benzoyl peroxide (BPO) 1.0 g
The mixed solution consisting of 30 g was dispersed in a solution consisting of 30 g of anhydrous sodium sulfate, 5 g of calcium carbonate, 20 ml of a 2% gelatin aqueous solution and 500 ml of water, and the temperature was raised from 28 ° C. to 60 ° C. in 1 hour. 60 ℃ for 3 hours, 70 ℃ for 1 hour, then 76
Polymerization was carried out at -80 ° C with stirring for 1 hour and 10 minutes.
After the toluene was distilled off under reduced pressure, 750 ml of water was added and the mixture was stirred at room temperature. 750 ml of water and 10 ml of concentrated sulfuric acid were added to the produced bead-like resin, and the mixture was stirred at room temperature for 30 minutes to decompose calcium carbonate, then washed successively with 750 ml of water and 300 ml of methanol and dried under reduced pressure. Acrylonitrile obtained-
The divinylbenzene copolymer was 95 g (yield 85.8).
%), And was in the form of light yellow beads.

Elemental analysis value (%): carbon 79.76, hydrogen 7.10, nitrogen 10.95.

Acrylonitrile / divinylbenzene ratio based on nitrogen: acrylonitrile / divinylbenzene = 0.64 / 0.36 (mol / mol) infrared absorption spectrum (cm ⁻¹ ): 2240 (-CN) 160
2 (1,4-substituted benzene).

(b) Acrylonitrile copolymer amidoximated hydroxyamine hydrochloride 6.75 g, sodium hydroxide 5.0
10.0 g of the acrylonitrile-divinylbenzene copolymer was added to a solution consisting of 9 g and 50 ml of methanol, and the mixture was stirred at 63 ° C. for 4 hours for reaction. After washing with methanol, it was dried under reduced pressure overnight to obtain a blood purification material having an amidoxime group as a side chain.

Example 2 (a) Synthesis of Acrylonitrile Grafted Cellulose While stirring a solution obtained by adding 16.20 g of cellulose powder and 15.89 g of acrylonitrile to 200 ml of water while stirring.
A solution prepared by dissolving 5 ml of a 1 M nitric acid solution of 0.1 M ceric ammonium nitrate was added, and the mixture was stirred at 24 to 30 ° C. for 2 hours for reaction. The reaction product was washed successively with water and methanol and then dried under reduced pressure at 50 ° C. for 24 hours to obtain acrylonitrile-grafted cellulose powder. Yield is 27.03g
Met. Elemental analysis values (%) were carbon 53.33, hydrogen 5.83, and nitrogen 11.09.

The above-mentioned acrylonitrile-grafted cellulose powder 10
g was extracted with a Soxhlet extractor using methanol as an extraction solvent at 7 minutes / cycle for 70 hours. Acrylonitrile grafted cellulose remaining as an extraction residue,
After further refluxing in methanol for washing, it was dried under reduced pressure. The yield was 7.3g. Elemental analysis values (%) were carbon 51.30, hydrogen 5.92, and nitrogen 9.62.

(b) Amidoximation of acrylonitrile-grafted cellulose, the acrylonitrile-grafted cellulose powder 1.
According to a conventional method, 50 g was reacted with hydroxyamine hydrochloride to obtain a blood purification material having an amidoxime group in its side chain.

The elemental analysis value (%) of the obtained blood purification material was 45.6 for carbon.
8, hydrogen was 6.24 and nitrogen was 8.86. Infrared spectrum (cm ^-1 ) is 1650 It was 2250 (-CN), but the peak corresponding to the nitrile group almost disappeared.

Example 3 5.0 g of acrylonitrile fiber (manufactured by Japan Exlan; 1.5 denier cut into 1 cm in length) was reacted with hydroxyamine hydrochloride according to a conventional method to prepare a blood purification material having an amidoxime group as a side chain. I got it.

The elemental analysis value (%) of the obtained blood purification material was 65.8 carbon.
3, hydrogen was 6.04, and nitrogen was 25.69. The amidoxime group calculated from the elemental analysis value was 0.93 meq / g.
Met.

(2) Production Example of Blood Purifying Material Having 2-Oxalohydrazinocarbonyl Group Example 4 (a) Synthesis of Etoxallylhydrazine A solution of 270 g (1.85 mol) of diethyl oxalate and 150 ml of ethanol was 90% hydrated. 66.7g hydrazine
A solution of (1.20 mol) in 75 ml of ethanol was added dropwise at -13 to -17 ° C over 30 minutes. Subsequently, after stirring at room temperature for 1 hour, the precipitate was filtered off, and ethanol and the like in the filtrate were distilled off at 30 ° C. and 5-35 mmHg. 300 ml of ethyl ethanol was added to the remaining oily substance and the mixture was left in the refrigerator overnight. The precipitated crystals were collected by filtration and recrystallized with ethyl ether to give etoxarylhydrazine crystals. The yield was 100.2 g (Y = 63%) and the melting point was 45-47 ° C.

The results of elemental analysis are as follows.

(b) Synthesis of 1-methacryloyl-ethoxalylhydrazine (abbreviation: MEH) 89.0 g of ethoxalylhydrazine (0.67mo
l), 67.8 g of triethylamine (0.67 mo)
1) and 850 ml of ethyl acetate while stirring, 70.5 g (0.67 m) of methacrylic acid chloride
ol) in 170 ml of ethyl acetate was added dropwise. The dropping was performed below freezing for 10 minutes. This solution
After further stirring at room temperature for 2 hours, the precipitate was filtered off, and the solvent was distilled off from the filtrate under reduced pressure to obtain crude MEH crystals. The crude crystals were extracted with ethyl ether using a Soxhlet extractor. The crystals precipitated from the ethyl ether layer of the extract were collected by filtration and dried under reduced pressure. The obtained crystals have a melting point of 10
The temperature was 3.5 to 104.5 ° C, and the yield was 70.3 g.

The elemental analysis is shown in the table below.

(c) Polymerization of MEH Homopolymerization of MEH and divinylbenzene (abbreviation: DV
The copolymerization with B) is carried out by azobisisobutyronitrile (AI
BN) was used as a catalyst.

The reaction conditions and the results are shown in Table 1.

The elemental analysis values of each polymer in Table 1 are as shown in Table 2.

(d) Hydrolysis of Polymer Polymer D2.0 was added with sodium hydroxide 4.0 g with water 10
After adding to a solution dissolved in 0 ml and stirring at 40 ° C. for 3 hours, the resin was collected by filtration. After washing with water, dipping in 0.2N hydrochloric acid for 2 hours, washing with water, and washing with methanol,
It was dried under reduced pressure at about 60 ° C for one day. Collection was 1.7 g. Elemental analysis values: carbon 50.80%, hydrogen 5.3
8%, nitrogen 11.01%, acid value 6.36 meq / g
Met. This hydrolyzed polymer is designated as F.

In addition, the polymers D and E were hydrolyzed under various conditions. The results are shown in Table 3.

(3) Production of Blood Purifying Material Containing Acid Hydrazide Group Example 5 35.7 m Methyl Methacrylate Purified by Vacuum Distillation
l, purity 55 washed with aqueous sodium hydroxide solution and water
% Divinylbenzene 1.88 ml, isooctane 18.
A solution consisting of 57 ml and 0.33 g of benzoyl peroxide was added to 15 ml of a 2% aqueous gelatin solution, 2 g of sodium sulfate,
The solution was dispersed in a solution of 2.35 g of calcium carbonate and 235 ml of water, and suspension polymerization was carried out at 70 to 75 ° C for 6 hours.
The purified polymerized product was separated by filtration, washed with water, diluted hydrochloric acid, water and then with hot water in that order, and then sieved to give a particle size of 0.42 to 0.42.
0.59 mm of the produced resin was collected. After being washed with hot water and hot methanol, dried under reduced pressure at 60 ° C. for a whole day and night to give resin 15.
6% was obtained. Elemental analysis values are carbon 61.1%, hydrogen 7.
It was 86%, and the divinylbenzene content determined based on the carbon analysis value was 2.9 mol%.

15 g of this resin and 75 ml of 90% hydrazine hydrate were charged into a glass cylindrical container and placed in an autoclave at 165 ° C.
The reaction was carried out by shaking for 7 hours. Then, after washing with water 10 times and then with hot methanol, it was dried under reduced pressure at 60 ° C. overnight to obtain 13.48 g of a resin. Elemental analysis value is carbon 5
0.20%, hydrogen 7.61%, nitrogen 21.99%, cation exchange capacity 2.45 meq / dry resin 1
g, anion exchange capacity was 4.35 meq / g of dry resin.

(4) Adsorption test of aluminum, pathogenic factors, etc. in blood Example 6 (Aluminum adsorption test) 5 ml of plasma collected from a long-term dialysis patient was mixed with 1 ml of the blood purification material of the present invention and shaken at 37 ° C. for 2 hours.・ After stirring, 30
Centrifugation was carried out at 00 rpm for 15 minutes, and the aluminum content of the supernatant was analyzed. The results are shown in Table 4.

From the results of Table 4, it was found that the blood purification material of the present invention well adsorbs aluminum in plasma.

Example 7 (Hepatitis B Antigen Adsorption Test) After weighing 0.5 g of the blood purification material of the present invention and putting it in a test tube having a volume of 6 ml, 1.5 ml of physiological saline was added to perform deaeration operation. What was defoamed after 2 hours was used for the experiment.

The hepatitis B antigen {HB
Add 1.5 ml of Ag (s, e) (+)} plasma, stir for 30 seconds, leave at 4 ° C for 1 day, and then add HBsAg and H
BeAg was measured by the RIA method and compared with untreated plasma. The result is displayed in titer value, It was expressed by the value of.

The results are shown in Table 5.

From this result, in the plasma containing the blood purification material of the present invention,
It was found that the hepatitis B antigen {HBAg (s, e) (+)} was almost removed by adsorption.

Example 8 (Adsorption test of 1 gG of pathogenic factor of multiple myeloid species) 0.5 mm of the blood purification material of the present invention was added to 1.5 ml of fresh human plasma, and the mixture was shaken and agitated at 37 ° C for 3 hours and then 3000 rpm.
Plasma was analyzed by centrifugation at. The results are shown in Table 6.

However, the upper numbers are before the adsorption test and the lower numbers are after the adsorption test.

The results of this test show that the blood purification material of the present invention adsorbs IgG, which is a pathogenic factor, well, but hardly adsorbs albumin, which is an essential protein.

Claims (3)

[Claims] 1. A blood purification material comprising a polymer having a functional group selected from the group consisting of an amidoxime group, a 2-oxahydrazide group and an acid hydrazide group in its side chain. 2. A method for removing aluminum in plasma, which comprises contacting plasma with the blood purification material according to claim 1. 3. A method for removing pathogenic factors and viruses in plasma, which comprises contacting plasma with the blood purification material according to claim 1.